Tumors express a range of antigens including self-antigens. CD4+CD25+ regulatory T (Treg) cells are critical for maintaining T cell tolerance to self-antigens. Treg cells are thought to dampen tumor associated antigen (TAA)-specific T cell immunity and to be the main obstacle tempering successful immunotherapy and active vaccination. Therefore, manipulation of regulatory T cells, including depletion, blocking trafficking into tumors, or reducing their differentiation/expansion, survival and suppressive mechanisms represent new strategies for cancer treatment. However, it remains poorly understood how Tregs survive and function in the environment with chronic hypoxia and nutrient depletion, and how the phenotypic and suppressor integrity of Tregs is maintained in the tumor microenvironment enriched with high levels of inflammatory factors. In this proposal, based on our preliminary data, we hypothesize that hypoxia activates the stemness program of Treg cells, reshapes local immune profile in the tumor microenvironment and contributes to tumor immune evasion. To test this hypothesis, extensive experiments are proposed herein along with three specific aims: Aim 1 is to test the hypothesis that hypoxia activates the stemness program in human tumor Tregs. Aim 2 is to test the hypothesis that hypoxia promotes Treg functional stability and integrity in human tumor. Aim 3 is to test the hypothesis that hypoxia activates key molecular targets in tumor Tregs.